sequences of Citrus junos
RNA-sequence and transcriptome analysis For transcriptome analysis, three biological replicates were used for diploid and tetraploid under well-watered or drought conditions. Total RNA was isolated from each treatment using the RNAplant Plus Reagent DP437 (Tiangen, Beijing, China) according to the manufacturers instructions. The concentration and quality of RNA were determined using an Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA, USA). Then, the RNA was sent to the Shanghai Personal Biotechnology Cp. Ltd. For RNA-Seq analysis (Personalbio, Shanghai, China). A total of 20-mg RNA was equally isolated from the each sample for cDNA library construction. Beads with oligo (dT) were used to enrich poly(A) mRNA, and then, fragmentation buffer was added for interrupting mRNA to short fragment (200~300 nt) which were used as templates to synthesize the first-strand cDNA using random hexamer primer, and the second-strand cDNA was synthesized by adding buffer, dNTPs, RNaseH, and DNA polymerase I, respectively. The doublestrand cDNA was purified with QiaQuick PCR Purification Kit (Qiagen, Germany) and washed with EB buffer for end repair and single nucleotide. Finally, the cDNA fragments were connected with sequencing adaptors and gel-electrophoresis was used to select cDNA fragments of 200 bp in size as the templates for amplification with PCR. The library products were ready for sequencing analysis via Illumina HiSeq 2000 (Illumina, San Diego, CA, USA). Bioinformatics analysis The RNA-Seq raw sequencing data was filtered using the Cutadapt program (http://cutadapt.readthedocs.io/en/stable/). The filtered clean reads were further mapped to the reference genome (HWB.chromosome.fa), downloaded from the database (http://citrus.hzau.edu.cn/orange/download/index.php) using Tophat2 (http://tophat.cbcb.umd.edu/), with up to two mismatches allowed. The transcripts were screened for significant changes in abundance using DESeq (http://www.bioconductor.org/packages/release/bioc/html/DESeq.html) with a conditio